Fig. 4

AZD1775 abrogates olaparib-activated DNA damage checkpoint and caused mitotic DNA damage in GC cells. Following cell cycle synchronization with serum deprivation for 24 h, 0.3 μM AZD1775 with/without 20 μM olaparib were exposed to MKN45 and AGS cells for 24 h. a and b Cell cycle was analyzed by flow cytometry. c Protein extracts were probed with anti-cyclin B1, CDC2 and pCDC2 antibodies. d and e pHH3 and γH2AX-positive cells at separate cell cycle stages were assessed by double-staining with pHH3/PI and γH2AX/PI, respectively, and quantified using FlowJo and ModFit software. AZD, AZD1775; Ola, olaparib; Com, Combination. Data expressed as Mean ± SD and representative of three independent experiments. *, P < 0.05 by ANOVA analysis